Correlation Between Polyphenol Oxidase (PPO) Activity and Total Phenolic Contents in Crocus sativus L. Corms During Dormancy and Sprouting Stages

Purification and characterization of polyphenol oxidase (PPO) enzyme and determination of total phenolic contents during dormancy and sprouting stages in Crocus sativus corms were performed. PPO enzyme was purified by ammonium sulfate precipitation and ion-exchange chromatography using DEAE-Sephadex A(25) and two isoenzymes were obtained on the SDS-PAGE, which corresponded to molecular weights of 70 and 54 kDa. The K(m) values of the enzyme were 4.87 and 2.12 mM for l-DOPA in dormancy and waking stages, respectively. Also, enzyme showed higher V(max) values of 0.026 (ΔOD.min(-1)) in dormancy compared with the value of 0.019 (ΔOD.min(-1)) in waking corms. Results showed an inverse correlation between phenolic contents and PPO activity. Accordingly, it can be concluded that as plant progressed through sprouting stage, in contrast to polyphenol oxidase activity, there was a significant increase in total amount of phenolic compounds, as determined by Folin-Ciocalteu method and water and aqueous ethanol extractions. SUMMARY: Purification of polyphenol oxidase enzyme using DEAE-Sephadex A(25) in Crocus sativus corms. Characterization of polyphenol oxidase enzyme. Comparison of PPO enzyme characteristics in two different physiologic stages of dormancy and sprouting. Determination of phenolic contents. Correlation between phenolic contents and PPO activity during sprouting and dormancy. [Image: see text] Abbreviations used: PPO: Polyphenol Oxidase, DEAE-Sephadex: Diethylaminoethyl Sephadex, SDS-PAGE: Sodium Dodecyl Sulfate- Polyacrylamide Gel Electrophoresis, DOPA: Dihydroxyphenylalanine, PEG: Polyethylene Glycol